LOCUS       sequence-191846-        7790 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LwCas13a locus into pACYC184 with nontargeting spacer.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7790)
  AUTHORS   Gootenberg JS, Abudayyeh OO, Lee JW, Essletzbichler P, Dy AJ, Joung 
            J, Verdine V, Donghia N, Daringer NM, Freije CA, Myhrvold C, 
            Bhattacharyya RP, Livny J, Regev A, Koonin EV, Hung DT, Sabeti PC, 
            Collins JJ, Zhang F
  TITLE     Nucleic acid detection with CRISPR-Cas13a/C2c2.
  JOURNAL   Science. 2017 Apr 13. pii: eaam9321. doi: 10.1126/science.aam9321.
  PUBMED    28408723
REFERENCE   2  (bases 1 to 7790)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2017 Apr 13. pii: eaam9321. doi: 10.1126/science.aam9321."
COMMENT     Sequence Label: pC012 LwCas13a locus with nontargeting spacer
FEATURES             Location/Qualifiers
     source          1..7790
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(154..188)
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     CDS             complement(192..3647)
                     /codon_start=1
                     /product="Leptotrichia wadei CRISPR-Cas effector that acts 
                     as an RNA-guided RNAse (Abudayyeh et al., 2017)"
                     /label=LwCas13a
                     /note="previously called C2c2"
                     /translation="MKVTKVDGISHKKYIEEGKLVKSTSEENRTSERLSELLSIRLDIY
                     IKNPDNASEEENRIRRENLKKFFSNKVLHLKDSVLYLKNRKEKNAVQDKNYSEEDISEY
                     DLKNKNSFSVLKKILLNEDVNSEELEIFRKDVEAKLNKINSLKYSFEENKANYQKINEN
                     NVEKVGGKSKRNIIYDYYRESAKRNDYINNVQEAFDKLYKKEDIEKLFFLIENSKKHEK
                     YKIREYYHKIIGRKNDKENFAKIIYEEIQNVNNIKELIEKIPDMSELKKSQVFYKYYLD
                     KEELNDKNIKYAFCHFVEIEMSQLLKNYVYKRLSNISNDKIKRIFEYQNLKKLIENKLL
                     NKLDTYVRNCGKYNYYLQVGEIATSDFIARNRQNEAFLRNIIGVSSVAYFSLRNILETE
                     NENDITGRMRGKTVKNNKGEEKYVSGEVDKIYNENKQNEVKENLKMFYSYDFNMDNKNE
                     IEDFFANIDEAISSIRHGIVHFNLELEGKDIFAFKNIAPSEISKKMFQNEINEKKLKLK
                     IFKQLNSANVFNYYEKDVIIKYLKNTKFNFVNKNIPFVPSFTKLYNKIEDLRNTLKFFW
                     SVPKDKEEKDAQIYLLKNIYYGEFLNKFVKNSKVFFKITNEVIKINKQRNQKTGHYKYQ
                     KFENIEKTVPVEYLAIIQSREMINNQDKEEKNTYIDFIQQIFLKGFIDYLNKNNLKYIE
                     SNNNNDNNDIFSKIKIKKDNKEKYDKILKNYEKHNRNKEIPHEINEFVREIKLGKILKY
                     TENLNMFYLILKLLNHKELTNLKGSLEKYQSANKEETFSDELELINLLNLDNNRVTEDF
                     ELEANEIGKFLDFNENKIKDRKELKKFDTNKIYFDGENIIKHRAFYNIKKYGMLNLLEK
                     IADKAKYKISLKELKEYSNKKNEIEKNYTMQQNLHRKYARPKKDEKFNDEDYKEYEKAI
                     GNIQKYTHLKNKVEFNELNLLQGLLLKILHRLVGYTSIWERDLRFRLKGEFPENHYIEE
                     IFNFDNSKNVKYKSGQIVEKYINFYKELYKDNVEKRSIYSDKKVKKLKQEKKDLYIRNY
                     IAHFNYIPHAEISLLEVLENLRKLLSYDRKLKNAIMKSIVDILKEYGFVATFKIGADKK
                     IEIQTLESEKIVHLKNLKKKKLMTDRNSEELCELVKVMFEYKALE"
     promoter        complement(join(3662..3667,3668..3685,3686..3691))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     rep_origin      3729..4274
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4175..4194
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4391..4408
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4800..4902
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             4903..5562
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4969..4988)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             complement(6596..7786)
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     7415..7434
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ggtgcctgac tgcgttagca atttaactgt gataaactac cgcattaaag cttgttttag
       61 tccccttcgt ttttggggta gtctaaatca gattgctgtt ctaccaagta atccatagtt
      121 ttagtcccct tcgtttttgg ggtagtctaa atcgctagca ttatacctag gactgagcta
      181 gctgtcaatc attccagggc cttgtactcg aacatcactt tcacgagttc gcacagttcc
      241 tcgctgttcc ggtcggtcat cagtttcttt ttcttcagat tcttcaggtg cacgatcttc
      301 tctgattcca gggtctggat ttcgatcttc ttgtcagcgc cgatcttgaa ggtggccacg
      361 aagccgtatt ctttcagaat gtccacgatg gacttcatga tggcgttctt cagcttccgg
      421 tcgtaggaca gcagcttccg caggttttcc agcacttcca gcaggctaat ctcggcgtgg
      481 gggatgtagt tgaagtgggc aatgtagttc cggatgtaca ggtccttttt ttcctgcttc
      541 agtttcttca ctttcttgtc ggagtagatg ctccgctttt ccacattgtc cttgtacagt
      601 tctttgtaga agttgatata cttttccacg atctggccgc ttttgtactt cacattcttg
      661 gagttgtcga aattgaaaat ttcctcgatg tagtggttct cgggaaactc gcccttcagc
      721 cggaatctca ggtcccgctc ccagatgctg gtgtagccca cgagccggtg caggatcttc
      781 agcagcaggc cctgcagcag gttcagctca ttgaattcca ccttgttctt caggtgggtg
      841 tacttctgga tgttgccgat ggccttctca tactctttgt agtcctcgtc gttgaacttt
      901 tcgtccttct tgggtctggc gtacttccgg tgcaggttct gctgcatggt gtagttcttt
      961 tcaatctcat tcttcttgtt gctgtactct ttcagttctt tcaggctgat cttatacttg
     1021 gccttatcgg cgatcttttc cagcagattc agcatgccgt atttcttgat attgtagaag
     1081 gcccggtgct tgatgatgtt ctcgccgtcg aaatagatct tgttggtgtc gaactttttc
     1141 agctctttcc ggtccttgat tttgttttcg ttgaagtcca ggaacttgcc gatctcgttg
     1201 gcttccagct cgaagtcctc ggtcactctg ttgttgtcca ggttcagcag gttgatcagt
     1261 tccagctcgt cgctgaaggt ttcttctttg ttggcggact ggtacttttc caggctgccc
     1321 ttcaggttgg tcagctcttt gtggttcagc agcttcagga tcaggtaaaa catgttcaga
     1381 ttctcggtgt acttcagaat cttccccagc ttgatctcgc gcacgaactc attgatctcg
     1441 tgagggattt ctttgttccg attgtgcttc tcatagttct tcaggatctt gtcgtacttc
     1501 tctttgttat cctttttgat cttgatcttg gagaagatgt cgttgttgtc attgttgttg
     1561 ttgctctcga tatacttcag attgttcttg ttcaggtagt cgatgaagcc cttcaggaaa
     1621 atctgctgaa taaagtcgat gtaggtattc ttttcctctt tgtcctggtt gttgatcatc
     1681 tctctgctct ggatgatggc caggtattcc acgggcacgg ttttctcgat gttctcgaac
     1741 ttctgatact tgtagtggcc ggttttctgg ttccgctgct tgttaatctt gatcacttca
     1801 ttggtgatct taaagaacac cttggagttt ttcacgaact tgttcaggaa ctcgccgtag
     1861 tagatattct tcagcaggta gatctgggcg tccttctctt ctttgtcctt gggcacgctc
     1921 caaaaaaact tcagggtatt ccgcaggtcc tcaatcttgt tgtacagctt ggtgaagctg
     1981 ggcacgaagg ggatgttttt gttcacgaag ttgaacttgg tattcttcag gtacttgatg
     2041 atcacatcct tctcgtagta gttgaacacg ttggcgctgt tcagctgctt gaagattttc
     2101 agcttcagct tcttttcgtt gatttcgttc tgaaacatct tcttggagat ctcgctgggg
     2161 gcgatattct tgaaggcgaa gatgtccttg ccttccagtt ccaggttgaa gtgcacgatg
     2221 ccgtgtctga tgctgctgat ggcctcgtcg atgttggcga agaagtcctc gatctcgttc
     2281 ttgttgtcca tgttgaagtc gtagctgtag aacatcttca gattttcttt cacttcgttc
     2341 tgcttgttct cattgtagat cttgtccacc tcgccggaca cgtatttctc ttcgcccttg
     2401 ttgttcttca cggtcttgcc ccgcatccgg ccggtgatat cgttctcgtt ctcggtttcc
     2461 aggatgttcc tcaggctgaa gtaggccacg ctggacacgc cgatgatgtt tctcaggaag
     2521 gcctcgttct gccggttccg ggcgataaag tcggaggtgg cgatctcgcc cacttgcaga
     2581 tagtagttgt acttgccgca gttccgcacg taggtgtcca gcttgttcag cagtttgttt
     2641 tcgatcagct ttttcagatt ctggtactcg aagatccgct tgatcttatc gttgctgatg
     2701 ttgctcagcc gcttgtacac gtagtttttc agcagctggg acatctcgat ttccacgaag
     2761 tggcagaagg cgtacttaat attcttgtcg ttcagttcct ctttgtccag gtagtacttg
     2821 tagaacacct ggcttttctt cagctcagac atgtcgggga tcttctcaat cagctctttg
     2881 atgttgttca cgttctggat ctcttcgtag ataatcttgg cgaagttctc tttgtcgttc
     2941 ttccggccga tgatcttgtg atagtactcg cggatcttgt acttctcgtg cttcttgctg
     3001 ttctcgatca ggaaaaacag tttctcgata tcctctttct tatacagctt gtcgaaggct
     3061 tcctgcacgt tgttgatgta gtcgttgcgc ttggcgctct ctctgtagta gtcgtagatg
     3121 atgttccgct tgctcttgcc gcccactttt tccacgttgt tctcgttgat cttctggtag
     3181 ttggccttgt tctcttcgaa gctgtacttc aggctgttga tcttgttcag cttggcttcc
     3241 acgtccttcc gaaagatttc cagttcctca gagttcacgt cctcgttcag caggatcttc
     3301 ttcagcacgg agaagctgtt cttgtttttc aggtcgtact cgctgatgtc ctcttcgcta
     3361 tagttcttgt cctgcacggc gttcttttct ttccggttct tcagatacag cacgctgtcc
     3421 ttcaggtgca gcaccttgtt gctaaagaac ttcttcaggt tctctcttct gatccggttc
     3481 tcttcctcgg aggcgttgtc ggggttcttg atgtagatgt ccagccggat gctcagcagc
     3541 tcgctcagtc tctcgctggt ccggttttcc tcgctggtgg acttcacgag cttgccctct
     3601 tcgatgtact tcttgtggct gatgccgtcg accttggtca ctttcatgga taagacctcc
     3661 taacatacga gccggaagca taaagtgtaa atctagaaat attttatctg attaataaga
     3721 tgatcttctt gagatcgttt tggtctgcgc gtaatctctt gctctgaaaa cgaaaaaacc
     3781 gccttgcagg gcggtttttc gaaggttctc tgagctacca actctttgaa ccgaggtaac
     3841 tggcttggag gagcgcagtc accaaaactt gtcctttcag tttagcctta accggcgcat
     3901 gacttcaaga ctaactcctc taaatcaatt accagtggct gctgccagtg gtgcttttgc
     3961 atgtctttcc gggttggact caagacgata gttaccggat aaggcgcagc ggtcggactg
     4021 aacggggggt tcgtgcatac agtccagctt ggagcgaact gcctacccgg aactgagtgt
     4081 caggcgtgga atgagacaaa cgcggccata acagcggaat gacaccggta aaccgaaagg
     4141 caggaacagg agagcgcacg agggagccgc cagggggaaa cgcctggtat ctttatagtc
     4201 ctgtcgggtt tcgccaccac tgatttgagc gtcagatttc gtgatgcttg tcaggggggc
     4261 ggagcctatg gaaaaacggc tttgccgcgg ccctctcact tccctgttaa gtatcttcct
     4321 ggcatcttcc aggaaatctc cgccccgttc gtaagccatt tccgctcgcc gcagtcgaac
     4381 gaccgagcgt agcgagtcag tgagcgagga agcggaatat atcctgtatc acatattctg
     4441 ctgacgcacc ggtgcagcct tttttctcct gccacatgaa gcacttcact gacaccctca
     4501 tcagtgccaa catagtaagc cagtatacac tccgctagcg ctgatgtccg gcggtgcttt
     4561 tgccgttacg caccaccccg tcagtagctg aacaggaggg acagctgata gaaacagaag
     4621 ccactggagc acctcaaaaa caccatcata cactaaatca gtaagttggc agcatcaccc
     4681 gacgcacttt gcgccgaata aatacctgtg acggaagatc acttcgcaga ataaataaat
     4741 cctggtgtcc ctgttgatac cgggaagccc tgggccaact tttggcgaaa atgacacgtt
     4801 gatcggcacg taagaggttc caactttcac cataatgaaa taagatcact accgggcgta
     4861 ttttttgagt tatcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact
     4921 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag
     4981 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag
     5041 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg
     5101 atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat
     5161 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg
     5221 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt
     5281 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtgtca
     5341 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc
     5401 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg
     5461 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat
     5521 gaattacaac agtactgcga tgagtggcag ggcggggcgt aattttttta aggcagttat
     5581 tggtgccctt aaacgcctgg ttgctacgcc tgaataagtg ataataagcg gatgaatggc
     5641 agaaattcga aagcaaattc gacccggtcg tcggttcagg gcagggtcgt taaatagccg
     5701 cttatgtcta ttgctggttt accggtttat tgactaccgg aagcagtgtg accgtgtgct
     5761 tctcaaatgc ctgaggccag tttgctcagg ctctccccgt ggaggtaata attgacgata
     5821 tgatcattta ttctgcctcc cagagcctga taaaaacggt tagcgcttcg ttaatacaga
     5881 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     5941 gcagggcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg
     6001 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     6061 gatagtcatg ccccgcgccc accggaagga gctaccggca gcggtgcgga ctgttgtaac
     6121 tcagaataag aaatgaggcc gctcatggcg ttgactctca gtcatagtat cgtggtatca
     6181 ccggttggtt ccactctctg ttgcgggcaa cttcagcagc acgtagggga cttccgcgtt
     6241 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
     6301 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
     6361 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
     6421 cgtggccagg acccaacgct gcccgagatg cgccgcgtgc ggctgctgga gatggcggac
     6481 gcgatggata tgttctgcca agggttggtt tgcgcattca cagttctccg caagaattga
     6541 ttggctccaa ttcttggagt ggtgaatccg ttagcgaggt gccgccggct tccattcagg
     6601 tcgaggtggc ccggctccat gcaccgcgac gcaacgcggg gaggcagaca aggtataggg
     6661 cggcgcctac aatccatgcc aacccgttcc atgtgctcgc cgaggcggca taaatcgccg
     6721 tgacgatcag cggtccaatg atcgaagtta ggctggtaag agccgcgagc gatccttgaa
     6781 gctgtccctg atggtcgtca tctacctgcc tggacagcat ggcctgcaac gcgggcatcc
     6841 cgatgccgcc ggaagcgaga agaatcataa tggggaaggc catccagcct cgcgtcgcga
     6901 acgccagcaa gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct
     6961 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     7021 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     7081 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     7141 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     7201 ctctcaaggg catcggtcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     7261 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     7321 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     7381 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     7441 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatccaca
     7501 ggacgggtgt ggtcgccatg atcgcgtagt cgatagtggc tccaagtagc gaagcgagca
     7561 ggactgggcg gcggccaaag cggtcggaca gtgctccgag aacgggtgcg catagaaatt
     7621 gcatcaacgc atatagcgct agcagcacgc catagtgact ggcgatgctg tcggaatgga
     7681 cgatatcccg caagaggccc ggcagtaccg gcataaccaa gcctatgcct acagcatcca
     7741 gggtgacggt gccgaggatg acgatgagcg cattgttaga tttcatacac
//
